Control mechanisms are of crucial importance for the maintenance of normal metabolism and a knowledge of the molecular details of the mechanisms that regulate metabolism is essential for elucidation of pathological processes. Thus, knowledge of the molecular mechanisms for regulation of the urea cycle, the major pathway of ammonia removal, will provide an understanding of Reye's syndrome and other disorders (e.g., hepatitis, cirrhosis, certain generalized disorders associated with viral and vascular disorders and metabolic defects) in which liver function is temporarily or permanently altered. Carbamyl phosphate synthetase, which catalyzes the first step of the urea cycle, is known to be the primary site of control, however, the molecular mechanisms for this control are not elucidated. Three mechanisms for the regulation of carbamyl phosphate synthetase activity have been detected in preliminary studies and the detailed operation of these mechanisms will be elucidated in the proposed research. Carbamyl phosphate synthetase is inhibited by physiological levels of Zn++ and this inhibition is reversed by an increased level of amino acids; equilibrium dialysis binding studies and atomic absorption spectrophotometry will be utilized to study this control mechanism. Carbamyl phosphate synthetase and ornithine transcarbamylase, which catalyzes the second step of the urea cycle, occur in extremely high concentrations in the mitochondrion. Self-association of carbamyl phosphate synthetase and complex formation between carbamyl phosphate synthetase and ornithine transcarbamylase also constitute regulatory mechanisms for the urea cycle. These mechanisms will be studied by performing sedimentation velocity, sedimentation equilibrium and covalent crosslinking experiments on the enzymes in solution; covalent crosslinking studies will also be used to define these protein-protein interactions within the intact mitochondrion.